In the past, immunoassays were developed for the quantitative and qualitative determination of a wide variety of compounds in a laboratory setting using detailed procedures and expensive instrumentation. Recent developments in immunodiagnostics have resulted in a movement toward more simple approaches to the rapid analysis of clinical samples. The development of solid phase bound reagents has eliminated the need for precipitation in the separation of bound reagents from free reagents. Further advancements in solid phase immunochemistry have resulted in non-instrumented dry reagent strip immunoassays. This configuration allows for the visual qualitative or semi-quantitative determination of analytes in patient samples without the use of an instrument.
There are two basic types of non-instrumented immunoassay configurations. In the first type, or visual color zone type, a signal is generated at a specific zone on the strip where the signal indicates the presence of analyte, and the intensity of the signal indicates the concentration of the analyte in the sample. This type of assay requires visual color interpretation either for the presence of color above a threshold, as in the case of a qualitative test, or the comparison of the color intensity to a color chart, as in the case of a semi-quantitative test. In the second type, the visual signal is produced along the length of a bibulous assay strip. During wicking, the analyte reacts with a signal-producing reagent and forms a visible signal along the support. The migration distance of the signal from the proximal end of the strip is a direct measure of analyte concentration. This type of non-instrumented migration height assay can achieve quantitative results with reasonable performance.
Although these single use, thermometer-type, non-instrumented quantitative devices and non-instrumented color comparison devices for qualitative measurement have shown adequate performance, they have several problems associated with reliability and convenience. The colors generated on these devices are not always uniform and sharp. In the case of migration type assays the border is often light in color, unclear and difficult to read. This translates directly into user errors since the user must make a judgment related to the position of the color band border. In the case of non-instrumented pregnancy tests it is sometimes difficult to visually interpret the intensity of the colored spot (especially at HCG concentrations close to the cut-off sensitivity), and interpretation of the result is sometimes a problem. Any time a non-technical operator is required to make a visual judgment or interpretation, an error is possible, and sometimes, is unavoidable.
To improve the accuracy and reliability of assay results, several qualitative and quantitative diagnostic tests have been developed in the clinical field utilizing a reflectometer for measuring optical radiation reflected from a test element such as a zone of detection on an analytical chemistry strip. Reflectometers have been constructed featuring optical arrangements of lenses, filters, apertures, a radiation source, and detector. Examples are described in U.S. Pat. Nos. 4,219,529, 4,224,032 and 3,536,927.
Often, problems arise with obtaining accurate and precise measurements of the diffusely reflected optical radiation from the surface of the assay strip within the sampling area or detection zone. One of the causes is that the physically detectable change which is being measured does not occur uniformly across the test area. A lateral flow assay strip passes a signal reagent through the detection zone which contains an immobilized capture reagent. The signal reagent first encounters and immediately begins binding with a substantial portion, if not the entire amount, of the capture reagent at the leading boundary of the detection zone. The result is a significantly higher signal intensity in a narrow region at the leading boundary, with a rapidly decreasing intensity towards the trailing boundary of the detection zone. Overall, the signal intensity is distributed in a non-uniform gradient across the detection zone.
Another potential cause of inaccurate measurements is high analyte concentrations which can produce a decrease in signal intensity or "hook effect." When the analyte concentration is significantly greater than the capture reagent concentration, the signal intensity can actually decrease, falsely indicating an acceptable assay result.
There is a need to achieve substantially uniform distribution of a physically detectable change across a detection zone using a capture reagent or other signal producing reagent. Another need is to achieve a predetermined distribution of the physically detectable change in the detection zone of an assay strip. These needs have not been filled by the prior art.
Thus, a need exists in the field of diagnostics for a method and device which varies the capture efficiency across a zone for detecting a physical change on the surface of a sample-exposed analytical chemistry strip. The uniform or predetermined distribution should be sufficiently inexpensive, timely, efficient, durable, and reliable for use in a diagnostic device which permits point-of-care use by untrained individuals in locations such as the home, sites of medical emergencies, or locations other than a clinic.